| 质粒名称: | pCRISPRyl_D17 |
|---|---|
| 出品公司: | Addgene |
| 目录编号: | 84611 |
| 存储实验室: | Ian Wheeldon |
|
载体骨架基本信息 |
|
| 载体名称: | pUC19 |
| 载体出品公司: | Clontech |
| 空载体大小: | 2623bp |
| 完整质粒大小: | 11822bp |
| 载体类型: | Yeast Expression, CRISPR, Synthetic Biology |
| 筛选标记: | LEU2 |
| 载体抗性: | 氨苄霉素 |
| 生长温度: | 37 ℃ |
| 宿主菌: | DH5a |
| 拷贝数: | 高拷贝 |
|
插入基因信息 |
|
| 插入基因名称: |
Cas9
gRNA/shRNA
|
| 启动子: | UAS1B8-TEF(136) |
| 标签/融合蛋白: | 人细胞色素c氧化酶亚基VIII（N端） |
| 克隆方法: | 限制性内切酶 |
| 5’克隆位点: | BssHII (not destroyed) |
| 3’克隆位点: | NheI (not destroyed) |
| 5’测序引物 | TEF |
| 3’测序引物 | SV40pA-R |
|
文献引用方法 |
|
| 材料和方法部分: | pCRISPRyl_D17 was a gift from Ian Wheeldon (Addgene plasmid # 84611 ; http://n2t.net/addgene:84611 ; RRID:Addgene_84611) |
| 参考文献部分: | Standardized markerless gene integration for pathway engineering in Yarrowia lipolytica. Schwartz C, Shabbir-Hussain M, Frogue K, Blenner M, Wheeldon I. ACS Synth Biol. 2016 Dec 19. 10.1021/acssynbio.6b00285 PubMed 27989123 |
买家导航
pCRISPRyl_D17载体质粒基本信息
pCRISPRyl_D17质粒图谱载体图谱和pCRISPRyl_D17载体序列质粒序列多克隆位点信息
pCRISPRyl_D17质粒载体简介
pCRISPRyl_D17质粒序列载体序列
LOCUS pCRISPRyl_D17 11823 bp DNA circular SYN 14-APR-2025
DEFINITION Introduces DSB at D17 locus in Y. lipolytica.
ACCESSION .
VERSION .
KEYWORDS .
SOURCE synthetic DNA construct
ORGANISM synthetic DNA construct
REFERENCE 1 (bases 1 to 11823)
AUTHORS Schwartz C, Shabbir-Hussain M, Frogue K, Blenner M, Wheeldon I
TITLE Standardized markerless gene integration for pathway engineering in
Yarrowia lipolytica.
JOURNAL ACS Synth Biol. 2016 Dec 19.
PUBMED 27989123
REFERENCE 2 (bases 1 to 11823)
AUTHORS .
TITLE Direct Submission
JOURNAL Exported Apr 14, 2025 from SnapGene Server 8.0.1
https://www.biofeng.com
COMMENT SGRef: number: 1; type: "Journal Article"; journalName: "ACS Synth
Biol. 2016 Dec 19."
COMMENT Sequence Label: pCRISPRyl_D17
FEATURES Location/Qualifiers
source 1..11823
/mol_type="other DNA"
/organism="synthetic DNA construct"
primer_bind complement(29..51)
/label=pGEX 3'
/note="pGEX vectors, reverse primer"
primer_bind 151..170
/label=pRS-marker
/note="pRS vectors, use to sequence yeast selectable
marker"
primer_bind 364..386
/label=M13/pUC Forward
/note="In lacZ gene"
primer_bind 378..395
/label=M13 Forward
/note="In lacZ gene. Also called M13-F20 or M13 (-21)
Forward"
primer_bind 379..395
/label=M13 fwd
/note="common sequencing primer, one of multiple similar
variants"
CDS 1740..5843
/codon_start=1
/product="Cas9 (Csn1) endonuclease from the Streptococcus
pyogenes Type II CRISPR/Cas system"
/label=Cas9
/note="generates RNA-guided double strand breaks in DNA"
/translation="MDKKYSIGLDIGTNSVGWAVITDEYKVPSKKFKVLGNTDRHSIKK
NLIGALLFDSGETAEATRLKRTARRRYTRRKNRICYLQEIFSNEMAKVDDSFFHRLEES
FLVEEDKKHERHPIFGNIVDEVAYHEKYPTIYHLRKKLVDSTDKADLRLIYLALAHMIK
FRGHFLIEGDLNPDNSDVDKLFIQLVQTYNQLFEENPINASGVDAKAILSARLSKSRRL
ENLIAQLPGEKKNGLFGNLIALSLGLTPNFKSNFDLAEDAKLQLSKDTYDDDLDNLLAQ
IGDQYADLFLAAKNLSDAILLSDILRVNTEITKAPLSASMIKRYDEHHQDLTLLKALVR
QQLPEKYKEIFFDQSKNGYAGYIDGGASQEEFYKFIKPILEKMDGTEELLVKLNREDLL
RKQRTFDNGSIPHQIHLGELHAILRRQEDFYPFLKDNREKIEKILTFRIPYYVGPLARG
NSRFAWMTRKSEETITPWNFEEVVDKGASAQSFIERMTNFDKNLPNEKVLPKHSLLYEY
FTVYNELTKVKYVTEGMRKPAFLSGEQKKAIVDLLFKTNRKVTVKQLKEDYFKKIECFD
SVEISGVEDRFNASLGTYHDLLKIIKDKDFLDNEENEDILEDIVLTLTLFEDREMIEER
LKTYAHLFDDKVMKQLKRRRYTGWGRLSRKLINGIRDKQSGKTILDFLKSDGFANRNFM
QLIHDDSLTFKEDIQKAQVSGQGDSLHEHIANLAGSPAIKKGILQTVKVVDELVKVMGR
HKPENIVIEMARENQTTQKGQKNSRERMKRIEEGIKELGSQILKEHPVENTQLQNEKLY
LYYLQNGRDMYVDQELDINRLSDYDVDHIVPQSFLKDDSIDNKVLTRSDKNRGKSDNVP
SEEVVKKMKNYWRQLLNAKLITQRKFDNLTKAERGGLSELDKAGFIKRQLVETRQITKH
VAQILDSRMNTKYDENDKLIREVKVITLKSKLVSDFRKDFQFYKVREINNYHHAHDAYL
NAVVGTALIKKYPKLESEFVYGDYKVYDVRKMIAKSEQEIGKATAKYFFYSNIMNFFKT
EITLANGEIRKRPLIETNGETGEIVWDKGRDFATVRKVLSMPQVNIVKKTEVQTGGFSK
ESILPKRNSDKLIARKKDWDPKKYGGFDSPTVAYSVLVVAKVEKGKSKKLKSVKELLGI
TIMERSSFEKNPIDFLEAKGYKEVKKDLIIKLPKYSLFELENGRKRMLASAGELQKGNE
LALPSKYVNFLYLASHYEKLKGSPEDNEQKQLFVEQHKHYLDEIIEQISEFSKRVILAD
ANLDKVLSAYNKHRDKPIREQAENIIHLFTLTNLGAPAAFKYFDTTIDRKRYTSTKEVL
DATLIHQSITGLYETRIDLSQLGGD"
CDS 1740..5843
/codon_start=1
/product="Cas9 (Csn1) endonuclease from the Streptococcus
pyogenes Type II CRISPR/Cas system"
/label=Cas9
/note="generates RNA-guided double strand breaks in DNA"
/translation="MDKKYSIGLDIGTNSVGWAVITDEYKVPSKKFKVLGNTDRHSIKK
NLIGALLFDSGETAEATRLKRTARRRYTRRKNRICYLQEIFSNEMAKVDDSFFHRLEES
FLVEEDKKHERHPIFGNIVDEVAYHEKYPTIYHLRKKLVDSTDKADLRLIYLALAHMIK
FRGHFLIEGDLNPDNSDVDKLFIQLVQTYNQLFEENPINASGVDAKAILSARLSKSRRL
ENLIAQLPGEKKNGLFGNLIALSLGLTPNFKSNFDLAEDAKLQLSKDTYDDDLDNLLAQ
IGDQYADLFLAAKNLSDAILLSDILRVNTEITKAPLSASMIKRYDEHHQDLTLLKALVR
QQLPEKYKEIFFDQSKNGYAGYIDGGASQEEFYKFIKPILEKMDGTEELLVKLNREDLL
RKQRTFDNGSIPHQIHLGELHAILRRQEDFYPFLKDNREKIEKILTFRIPYYVGPLARG
NSRFAWMTRKSEETITPWNFEEVVDKGASAQSFIERMTNFDKNLPNEKVLPKHSLLYEY
FTVYNELTKVKYVTEGMRKPAFLSGEQKKAIVDLLFKTNRKVTVKQLKEDYFKKIECFD
SVEISGVEDRFNASLGTYHDLLKIIKDKDFLDNEENEDILEDIVLTLTLFEDREMIEER
LKTYAHLFDDKVMKQLKRRRYTGWGRLSRKLINGIRDKQSGKTILDFLKSDGFANRNFM
QLIHDDSLTFKEDIQKAQVSGQGDSLHEHIANLAGSPAIKKGILQTVKVVDELVKVMGR
HKPENIVIEMARENQTTQKGQKNSRERMKRIEEGIKELGSQILKEHPVENTQLQNEKLY
LYYLQNGRDMYVDQELDINRLSDYDVDHIVPQSFLKDDSIDNKVLTRSDKNRGKSDNVP
SEEVVKKMKNYWRQLLNAKLITQRKFDNLTKAERGGLSELDKAGFIKRQLVETRQITKH
VAQILDSRMNTKYDENDKLIREVKVITLKSKLVSDFRKDFQFYKVREINNYHHAHDAYL
NAVVGTALIKKYPKLESEFVYGDYKVYDVRKMIAKSEQEIGKATAKYFFYSNIMNFFKT
EITLANGEIRKRPLIETNGETGEIVWDKGRDFATVRKVLSMPQVNIVKKTEVQTGGFSK
ESILPKRNSDKLIARKKDWDPKKYGGFDSPTVAYSVLVVAKVEKGKSKKLKSVKELLGI
TIMERSSFEKNPIDFLEAKGYKEVKKDLIIKLPKYSLFELENGRKRMLASAGELQKGNE
LALPSKYVNFLYLASHYEKLKGSPEDNEQKQLFVEQHKHYLDEIIEQISEFSKRVILAD
ANLDKVLSAYNKHRDKPIREQAENIIHLFTLTNLGAPAAFKYFDTTIDRKRYTSTKEVL
DATLIHQSITGLYETRIDLSQLGGD"
CDS 5856..5876
/codon_start=1
/product="nuclear localization signal of SV40 (simian virus
40) large T antigen"
/label=SV40 NLS
/translation="PKKKRKV"
terminator 5887..6134
/gene="S. cerevisiae CYC1"
/label=CYC1 terminator
/note="transcription terminator for CYC1"
primer_bind complement(5903..5921)
/label=CYC1
/note="CYC1 transcription termination signal, reverse
primer"
primer_bind complement(6153..6169)
/label=M13 rev
/note="common sequencing primer, one of multiple similar
variants"
primer_bind complement(6153..6169)
/label=M13 Reverse
/note="In lacZ gene. Also called M13-rev"
primer_bind complement(6166..6188)
/label=M13/pUC Reverse
/note="In lacZ gene"
protein_bind 6177..6193
/label=lac operator
/bound_moiety="lac repressor encoded by lacI"
/note="The lac repressor binds to the lac operator to
inhibit transcription in E. coli. This inhibition can be
relieved by adding lactose or
isopropyl-beta-D-thiogalactopyranoside (IPTG)."
promoter complement(join(6201..6207,6208..6225,6226..6231))
/label=lac promoter
/note="promoter for the E. coli lac operon"
protein_bind 6246..6267
/label=CAP binding site
/bound_moiety="E. coli catabolite activator protein"
/note="CAP binding activates transcription in the presence
of cAMP."
primer_bind complement(6384..6401)
/label=L4440
/note="L4440 vector, forward primer"
rep_origin complement(6555..7143)
/direction=LEFT
/label=ori
/note="high-copy-number ColE1/pMB1/pBR322/pUC origin of
replication"
primer_bind complement(6635..6654)
/label=pBR322ori-F
/note="pBR322 origin, forward primer"
CDS complement(join(7314..8105,8106..8174))
/codon_start=1
/gene="bla"
/product="beta-lactamase"
/label=AmpR
/note="confers resistance to ampicillin, carbenicillin, and
related antibiotics"
/translation="MSIQHFRVALIPFFAAFCLPVFAHPETLVKVKDAEDQLGARVGYI
ELDLNSGKILESFRPEERFPMMSTFKVLLCGAVLSRIDAGQEQLGRRIHYSQNDLVEYS
PVTEKHLTDGMTVRELCSAAITMSDNTAANLLLTTIGGPKELTAFLHNMGDHVTRLDRW
EPELNEAIPNDERDTTMPVAMATTLRKLLTGELLTLASRQQLIDWMEADKVAGPLLRSA
LPAGWFIADKSGAGERGSRGIIAALGPDGKPSRIVVIYTTGSQATMDERNRQIAEIGAS
LIKHW"
primer_bind 7937..7956
/label=Amp-R
/note="Ampicillin resistance gene, reverse primer"
promoter complement(8175..8278)
/gene="bla"
/label=AmpR promoter
misc_RNA 8773..8848
/label=gRNA scaffold
/note="guide RNA scaffold for the Streptococcus pyogenes
CRISPR/Cas9 system"
primer_bind 8898..8916
/label=pBRforEco
/note="pBR322 vectors, upsteam of EcoRI site, forward
primer"
ORIGIN
1 tcgcgcgttt cggtgatgac ggtgaaaacc tctgacacat gcagctcccg gagacggtca
61 cagcttgtct gtaagcggat gccgggagca gacaagcccg tcagggcgcg tcagcgggtg
121 ttggcgggtg tcggggctgg cttaactatg cggcatcaga gcagattgta ctgagagtgc
181 accatatgcg gtgtgaaata ccgcacagat gcgtaaggag aaaataccgc atcaggcgcc
241 attcgccatt caggctgcgc aactgttggg aagggcgatc ggtgcgggcc tcttcgctat
301 tacgccagct ggcgaaaggg ggatgtgctg caaggcgatt aagttgggta acgccagggt
361 tttcccagtc acgacgttgt aaaacgacgg ccagtgaatt cgagctcggt acccgctttt
421 cgtagataat ggaatacaaa tggatatcca gagtatacac atggatagta tacactgaca
481 cgacaattct gtatctcttt atgttaacta ctgtgaggcg ttaaatagag cttgatatat
541 aaaatgttac atttcacagt ctgaactttt gcagattacc taatttggta agatattaat
601 tatgaactga aagttgatgg catccctaaa tttgatgaaa gggggatccc ccgggttcga
661 aggtaccaag gaagcatgcg gtacccgaat tcctgaggtg tctcacaagt gccgtgcagt
721 cccgccccca cttgcttctc tttgtgtgta gtgtacgtac attatcgaga ccgttgttcc
781 cgcccacctc gatccggtct agactgaggt gtctcacaag tgccgtgcag tcccgccccc
841 acttgcttct ctttgtgtgt agtgtacgta cattatcgag accgttgttc ccgcccacct
901 cgatccgggg atccctgagg tgtctcacaa gtgccgtgca gtcccgcccc cacttgcttc
961 tctttgtgtg tagtgtacgt acattatcga gaccgttgtt cccgcccacc tcgatccggg
1021 tcgacctgag gtgtctcaca agtgccgtgc agtcccgccc ccacttgctt ctctttgtgt
1081 gtagtgtacg tacattatcg agaccgttgt tcccgcccac ctcgatccgg gagctcctga
1141 ggtgtctcac aagtgccgtg cagtcccgcc cccacttgct tctctttgtg tgtagtgtac
1201 gtacattatc gagaccgttg ttcccgccca cctcgatccg gtctagactg aggtgtctca
1261 caagtgccgt gcagtcccgc ccccacttgc ttctctttgt gtgtagtgta cgtacattat
1321 cgagaccgtt gttcccgccc acctcgatcc ggggatccct gaggtgtctc acaagtgccg
1381 tgcagtcccg cccccacttg cttctctttg tgtgtagtgt acgtacatta tcgagaccgt
1441 tgttcccgcc cacctcgatc cgggtcgacc tgaggtgtct cacaagtgcc gtgcagtccc
1501 gcccccactt gcttctcttt gtgtgtagtg tacgtacatt atcgagaccg ttgttcccgc
1561 ccacctcgat ccgggcatgc ctgcagaagc ttttgtggtt gggactttag ccaagggtat
1621 aaaagaccac cgtccccgaa ttacctttcc tcttcttttc tctctctcct tgtcaactca
1681 cacccgaaat cgttaagcat ttccttctga gtataagaat cattcaaagg cgcgccacaa
1741 tggataagaa atactccatt ggcctggaca tcggaaccaa ctccgtgggt tgggccgtga
1801 tcaccgatga gtacaaggtg ccctctaaga aattcaaggt cctgggcaac accgaccgac
1861 actccatcaa gaagaacctg atcggcgctc tgctcttcga ctctggcgag accgctgagg
1921 ccacccgact gaagcgaacc gctcgaagac gatacacccg aagaaagaac cgaatctgtt
1981 acctgcagga gatcttctct aacgagatgg ccaaggtgga cgactctttc ttccaccgac
2041 tggaggagtc tttcctggtg gaggaggaca agaagcacga gcgacacccc atcttcggca
2101 acatcgtgga cgaggtggcc taccacgaga agtaccccac catctaccac ctgcgaaaga
2161 agctggtgga ctctaccgac aaggccgacc tgcgactgat ctacctggcc ctggcccaca
2221 tgatcaagtt ccgaggccac ttcctgatcg agggcgacct gaaccccgac aactctgacg
2281 tggacaagct gttcatccag ctggtgcaga cctacaacca gctcttcgaa gagaacccca
2341 ttaacgcttc tggcgtggat gctaaggcca tcctgtctgc ccgactgtct aagtctcgac
2401 gactcgagaa cctgattgct cagctccccg gagagaagaa gaacggtctg ttcggaaacc
2461 tgattgctct gtccctgggt ctcaccccta acttcaagtc caacttcgat ctggctgagg
2521 acgctaagct gcagctgtct aaggacacct acgacgatga cctggataac ctgctcgccc
2581 agattggcga ccagtacgcc gacctgttcc tggccgccaa gaacctgtct gacgccatcc
2641 tgctgtctga catcctgcga gtgaacaccg agatcaccaa ggcccccctg tctgcctcca
2701 tgattaagcg atacgatgag caccaccagg atctgaccct cctcaaggct ctggtccgac
2761 agcagctgcc cgagaagtac aaggagattt tcttcgacca gtctaagaac ggctacgccg
2821 gctacatcga cggcggcgcc tctcaggagg agttctacaa gttcattaag cccatcctgg
2881 agaagatgga cggaaccgag gaactgctcg tgaagctgaa ccgagaggac ctcctgcgaa
2941 agcagcgaac cttcgacaac ggctctatcc cccaccagat ccacctgggc gagctgcacg
3001 ccatcctgcg acgacaggag gacttctacc ccttcctgaa ggacaaccga gagaagatcg
3061 agaagatcct gaccttccga atcccctact acgtgggacc cctggcccga ggaaactctc
3121 gattcgcttg gatgacccga aagtctgagg agaccattac cccctggaac ttcgaggagg
3181 tggtggataa gggcgcctct gctcagtctt tcatcgagcg aatgaccaac ttcgacaaga
3241 acctccccaa cgagaaggtc ctgcccaagc actctctgct ctacgagtac ttcaccgtct
3301 acaacgagct caccaaggtc aagtacgtga ccgagggaat gcgaaagccc gctttcctgt
3361 ctggagagca gaagaaggct attgtggatc tgctcttcaa gactaaccga aaggtcaccg
3421 tcaagcagct gaaggaggat tacttcaaga agattgagtg tttcgattct gtcgagatct
3481 ccggcgtcga ggaccgattc aacgcctctc tgggtaccta ccacgacctg ctgaagatta
3541 tcaaggacaa ggatttcctg gataacgagg agaacgagga tattctcgag gacattgtcc
3601 tgaccctcac cctgttcgag gatcgagaga tgattgagga gcgactcaag acctacgctc
3661 acctgttcga cgacaaggtg atgaagcagc tgaagcgacg acgatacacc ggctggggcc
3721 gactgtctcg aaagctgatc aacggcatcc gagacaagca gtctggcaag accatcctgg
3781 acttcctgaa gtctgacggc ttcgccaacc gaaacttcat gcagctgatc cacgacgact
3841 ctctgacctt caaggaggac atccagaagg cccaggtgtc tggccagggc gactctctgc
3901 acgagcacat cgccaacctg gccggctctc ccgccattaa gaaaggtatc ctgcagaccg
3961 tcaaggtggt cgatgagctc gtcaaggtga tgggccgaca caagcccgag aacattgtca
4021 ttgagatggc tcgagagaac cagactactc agaagggtca gaaaaactcc cgagagcgaa
4081 tgaagcgaat tgaggaaggt attaaggagc tgggatccca gattctcaag gagcatcccg
4141 tggagaacac tcagctccag aacgagaagc tgtacctgta ctatctgcag aacggtcgag
4201 acatgtacgt cgaccaggag ctggatatca accgactctc cgactacgat gtggaccaca
4261 ttgtgcccca gtccttcctg aaggacgatt ctatcgataa caaggtgctg acccgatccg
4321 acaagaaccg aggcaagtct gacaacgtgc cctccgagga ggtggtcaag aagatgaaga
4381 actactggcg acagctgctg aacgccaagc tgattaccca gcgaaagttc gacaacctga
4441 ccaaggccga gcgaggcggc ctgtctgagc tggacaaggc cggcttcatc aagcgacagc
4501 tggtggagac ccgacagatc accaagcacg tggcccagat cctggactct cgaatgaaca
4561 ccaagtacga cgagaacgac aagctgatcc gagaggtgaa ggtgatcacc ctgaagtcta
4621 agctggtgtc tgacttccga aaggacttcc agttctacaa ggtgcgagag attaacaact
4681 accaccacgc ccacgatgcc tacctgaacg ctgtcgtggg caccgccctc atcaagaagt
4741 atcccaagct ggagtccgag ttcgtctacg gcgactacaa ggtctacgat gtgcgaaaaa
4801 tgattgccaa gtccgagcag gagattggca aggctaccgc caagtacttc ttctactcca
4861 acattatgaa cttcttcaag accgagatta ccctggctaa cggcgagatt cgaaagcgac
4921 ccctcattga gaccaacgga gagaccggtg agatcgtgtg ggacaaggga cgagacttcg
4981 ccaccgtgcg aaaggtgctg tctatgcccc aggtgaacat cgtgaagaag accgaggtgc
5041 agaccggagg tttctctaag gagtccatcc tgcccaagcg aaactctgac aagctgatcg
5101 cccgaaagaa ggactgggac cccaagaagt acggaggttt cgactctccc accgtggctt
5161 actctgtgct ggtggtggcc aaggtggaga agggcaagtc taagaagctg aagtctgtga
5221 aggagctgct gggcattacc atcatggagc gatcttcttt cgagaagaac cccattgact
5281 tcctggaggc caagggatac aaggaggtga agaaagatct gattatcaag ctccccaagt
5341 actctctgtt cgagctggag aacggacgaa agcgaatgct ggcctctgcc ggcgagctgc
5401 agaagggaaa cgagctggcc ctgccctcca agtacgtcaa cttcctgtac ctcgcctccc
5461 attacgagaa gctgaagggc tctcccgagg ataacgagca gaagcagctc ttcgtggagc
5521 agcataagca ctacctggac gagatcatcg agcagatctc tgagttctct aagcgagtga
5581 tcctggctga cgccaacctg gataaggtgc tgtctgctta caacaagcac cgagacaagc
5641 ccattcgaga gcaggctgag aacatcattc acctgttcac cctgaccaac ctgggagccc
5701 ccgctgcctt caagtacttc gacaccacca tcgaccgaaa gcgatacacc tctaccaagg
5761 aggtgctgga cgccaccctg atccaccagt ctatcaccgg cctgtacgag acccgaatcg
5821 acctgtctca gctgggcggc gactctcgag ccgaccccaa gaagaagcga aaggtgtaag
5881 ctagcctcat gtaattagtt atgtcacgct tacattcacg ccctcccccc acatccgctc
5941 taaccgaaaa ggaaggagtt agacaacctg aagtctaggt ccctatttat ttttttatag
6001 ttatgttagt attaagaacg ttatttatat ttcaaatttt tctttttttt ctgtacagac
6061 gcgtgtacgc atgtaacatt atactgaaaa ccttgcttga gaaggttttg ggacgctcga
6121 aggctttaat ttgcaagctt ggcgtaatca tggtcatagc tgtttcctgt gtgaaattgt
6181 tatccgctca caattccaca caacatacga gccggaagca taaagtgtaa agcctggggt
6241 gcctaatgag tgagctaact cacattaatt gcgttgcgct cactgcccgc tttccagtcg
6301 ggaaacctgt cgtgccagct gcattaatga atcggccaac gcgcggggag aggcggtttg
6361 cgtattgggc gctcttccgc ttcctcgctc actgactcgc tgcgctcggt cgttcggctg
6421 cggcgagcgg tatcagctca ctcaaaggcg gtaatacggt tatccacaga atcaggggat
6481 aacgcaggaa agaacatgtg agcaaaaggc cagcaaaagg ccaggaaccg taaaaaggcc
6541 gcgttgctgg cgtttttcca taggctccgc ccccctgacg agcatcacaa aaatcgacgc
6601 tcaagtcaga ggtggcgaaa cccgacagga ctataaagat accaggcgtt tccccctgga
6661 agctccctcg tgcgctctcc tgttccgacc ctgccgctta ccggatacct gtccgccttt
6721 ctcccttcgg gaagcgtggc gctttctcaa tgctcacgct gtaggtatct cagttcggtg
6781 taggtcgttc gctccaagct gggctgtgtg cacgaacccc ccgttcagcc cgaccgctgc
6841 gccttatccg gtaactatcg tcttgagtcc aacccggtaa gacacgactt atcgccactg
6901 gcagcagcca ctggtaacag gattagcaga gcgaggtatg taggcggtgc tacagagttc
6961 ttgaagtggt ggcctaacta cggctacact agaaggacag tatttggtat ctgcgctctg
7021 ctgaagccag ttaccttcgg aaaaagagtt ggtagctctt gatccggcaa acaaaccacc
7081 gctggtagcg gtggtttttt tgtttgcaag cagcagatta cgcgcagaaa aaaaggatct
7141 caagaagatc ctttgatctt ttctacgggg tctgacgctc agtggaacga aaactcacgt
7201 taagggattt tggtcatgag attatcaaaa aggatcttca cctagatcct tttaaattaa
7261 aaatgaagtt ttaaatcaat ctaaagtata tatgagtaaa cttggtctga cagttaccaa
7321 tgcttaatca gtgaggcacc tatctcagcg atctgtctat ttcgttcatc catagttgcc
7381 tgactccccg tcgtgtagat aactacgata cgggagggct taccatctgg ccccagtgct
7441 gcaatgatac cgcgagaccc acgctcaccg gctccagatt tatcagcaat aaaccagcca
7501 gccggaaggg ccgagcgcag aagtggtcct gcaactttat ccgcctccat ccagtctatt
7561 aattgttgcc gggaagctag agtaagtagt tcgccagtta atagtttgcg caacgttgtt
7621 gccattgcta caggcatcgt ggtgtcacgc tcgtcgtttg gtatggcttc attcagctcc
7681 ggttcccaac gatcaaggcg agttacatga tcccccatgt tgtgcaaaaa agcggttagc
7741 tccttcggtc ctccgatcgt tgtcagaagt aagttggccg cagtgttatc actcatggtt
7801 atggcagcac tgcataattc tcttactgtc atgccatccg taagatgctt ttctgtgact
7861 ggtgagtact caaccaagtc attctgagaa tagtgtatgc ggcgaccgag ttgctcttgc
7921 ccggcgtcaa tacgggataa taccgcgcca catagcagaa ctttaaaagt gctcatcatt
7981 ggaaaacgtt cttcggggcg aaaactctca aggatcttac cgctgttgag atccagttcg
8041 atgtaaccca ctcgtgcacc caactgatct tcagcatctt ttactttcac cagcgtttct
8101 gggtgagcaa aaacaggaag gcaaaatgcc gcaaaaaagg gaataagggc gacacggaaa
8161 tgttgaatac tcatactctt cctttttcaa tattattgaa gcatttatca gggttattgt
8221 ctcatgagcg gatacatatt tgaatgtatt tagaaaaata aacaaatagg ggttccgcac
8281 acatttcccc gaaaagtgcc acctgacgtc cccagttgca aaagttgaca caactctaga
8341 tctgcttcca aatatagaat cataacaagg gttagggtgt gattatataa tattggtctt
8401 aattgatgtg ctagggcttt aaaagttggt taaaataacg ctctaatgcc tttttaatat
8461 attgtctttt tcaaaatctc aaatcggaca cttcttcgtg tatgagactc cattttttgg
8521 ctccgtcacg tgatatgtat tatcagctat agtggtgtaa acaaagtttt ttactagctg
8581 taatggcatt ttgtcggagt ggtaaatcgc cttcttgttg tgcgttcgag ttctggactc
8641 tgcactgggc tactttgaaa aatacctcta atgcgccgat ggtttagtgg taaaatccat
8701 cgttgccatc gatgggcccc cggttcgatt ccgggtcggc gcaggttgac gttccgtaat
8761 ataggtgacg acgttttaga gctagaaata gcaagttaaa ataaggctag tccgttatca
8821 acttgaaaaa gtggcaccga gtcggtgctt ttttttacgt ctaagaaacc attattatca
8881 tgacattaac ctataaaaat aggcgtatca cgaggcccag atcctctaga gtcgaagcgg
8941 ccgctatgtc tgataaaagg atgtaacata ggcaagctgc tcgtgagtgt tgagtacgaa
9001 ccttagatcc aaatcacccg cacccacgga tatacttgct tgaatataca gtagtatgcg
9061 gccgcttcga caccatatca tataaaacta acaatgcatt gaattaatac gaagactacc
9121 cgttgctatc tccacaccgt tatctccacg gtccaaaggc tgctcaatgt gctgcatacg
9181 taacgtgggg tgcaaccttg agcacatagt acttttccga aaaccggcga taattaagtg
9241 tgcactccaa cttttcacac tgagcgtaaa atgtggagaa gaaatcggca ctaaaaagtc
9301 aggtagactg gaaaatgcgc catgaaatga atatctcttg ctacagtaat gcccagcatc
9361 gaggggtatt gtgtcaccaa cactatagtg gcagctgaag cgctcgtgat tgtagtatga
9421 gtctttattg gtgatgggaa gagttcactc aatattctcg ttactgccaa aacaccacgg
9481 taatcggcca gacaccatgg atgtagatca ccaagcctgt gaatgttatt cgagctaaaa
9541 tgcacatggt tggtgaaagg agtagttgct gtcgaattcc gtcgtcgcct gagtcatcat
9601 ttatttacca gttggccaca aacccttgac gatctcgtat gtcccctccg acatactccc
9661 ggccggctgg ggtacgttcg atagcgctat cggcatcgac aaggtttggg tccctagccg
9721 ataccgcact acctgagtca caatcttcgg aggtttagtc ttccacatag cacgggcaaa
9781 agtgcgtata tatacaagag cgtttgccag ccacagattt tcactccaca caccacatca
9841 cacatacaac cacacacatc cacaatggaa cccgaaacta agaagaccaa gactgactcc
9901 aagaagattg ttcttctcgg cggcgacttc tgtggccccg aggtgattgc cgaggccgtc
9961 aaggtgctca agtctgttgc tgaggcctcc ggcaccgagt ttgtgtttga ggaccgactc
10021 attggaggag ctgccattga gaaggagggc gagcccatca ccgacgctac tctcgacatc
10081 tgccgaaagg ctgactctat tatgctcggt gctgtcggag gcgctgccaa caccgtatgg
10141 accactcccg acggacgaac cgacgtgcga cccgagcagg gtctcctcaa gctgcgaaag
10201 gacctgaacc tgtacgccaa cctgcgaccc tgccagctgc tgtcgcccaa gctcgccgat
10261 ctctccccca tccgaaacgt tgagggcacc gacttcatca ttgtccgaga gctcgtcgga
10321 ggtatctact ttggagagcg aaaggaggat gacggatctg gcgtcgcttc cgacaccgag
10381 acctactccg ttcctgaggt tgagcgaatt gcccgaatgg ccgccttcct ggcccttcag
10441 cacaaccccc ctcttcccgt gtggtctctt gacaaggcca acgtgctggc ctcctctcga
10501 ctttggcgaa agactgtcac tcgagtcctc aaggacgaat tcccccagct cgagctcaac
10561 caccagctga tcgactcggc cgccatgatc ctcatcaagc agccctccaa gatgaatggt
10621 atcatcatca ccaccaacat gtttggcgat atcatctccg acgaggcctc cgtcatcccc
10681 ggttctctgg gtctgctgcc ctccgcctct ctggcttctc tgcccgacac caacgaggcg
10741 ttcggtctgt acgagccctg tcacggatct gcccccgatc tcggcaagca gaaggtcaac
10801 cccattgcca ccattctgtc tgccgccatg atgctcaagt tctctcttaa catgaagccc
10861 gccggtgacg ctgttgaggc tgccgtcaag gagtccgtcg aggctggtat cactaccgcc
10921 gatatcggag gctcttcctc cacctccgag gtcggagact tgttgccaac aaggtcaagg
10981 agctgctcaa gaaggagtaa gtcgtttcta cgacgcattg atggaaggag caaactgacg
11041 cgcctgcggg ttggtctacc ggcagggtcc gctagtgtat aagactctat aaaaagggcc
11101 ctgccctgct aatgaaatga tgatttataa tttaccggtg tagcaacctt gactagaaga
11161 agcagattgg gtgtgtttgt agtggaggac agtggtacgt tttggaaaca gtcttcttga
11221 aagtgtcttg tctacagtat attcactcat aacctcaata gccaagggtg tagtcggttt
11281 attaaaggaa gggagttgtg gctgatgtgg atagatatct ttaagctggc gactgcaccc
11341 aacgagtgtg gtggtagctt gttactgtat attcggtaag atatattttg tggggtttta
11401 gtggtgtttg gtaggttagt gcttggtata tgagttgtag gcatgacaat ttggaaaggg
11461 gtggactttg ggaatattgt gggatttcaa taccttagtt tgtacagggt aattgttaca
11521 aatgatacaa agaactgtat ttcttttcat ttgttttaat tggttgtata tcaagtccgt
11581 tagacgagct cagtgccatg gcttttggca ctgtatttca tttttagagg tacactacat
11641 tcagtgaggt atggtaaggt tgagggcata atgaaggcac cttgtactga cagtcacaga
11701 cctctcaccg agaattttat gagatatact cgggttcatt ttaggctccg attcgattca
11761 aattattact gtcgaaatcg gttgagcatc cgttgatttc cgaacagatc tggccctttc
11821 gtc
//
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